Functional analysis of RNA polymerase II Rpb3 mutants of the fission yeast Schizosaccharomyces pombe.

Abstract:

:The RNA polymerase II (Pol II) of Schizosaccharomyces pombe is composed of 12 subunits. Subunit Rpb3 has sequence homology with the N-terminal domain of the prokaryotic alpha subunit, which plays a key role in RNA polymerase assembly. Together with the Rpb2 (the beta homologue) and Rpb11 (the second alpha homologue) subunits, Rpb3 constitutes a core subassembly (Rpb2-Rpb3-Rpb11) which corresponds to the the alpha2beta assembly intermediate of prokaryotic RNA polymerase. For the functional mapping of Rpb3, we made a collection of 12 heat-sensitive (Ts) or cold-sensitive (Cs) S. pombe mutants, each carrying a single mutation in one of the four conserved regions of Rpb3. The altered functions of six representative Pol II mutants containing the mutant Rpb3 were analyzed in vitro using an improved version of the GAL4-VP16 activator-dependent transcription system catalyzed by S. pombe cell extracts. The transcription activity by the extracts from Rpb3 mutants decreased to varying extents after heat treatment; but the extracts from Rpb3 mutants which had mutations in the eukaryote-specific conserved regions B and C regained their activity by the addition of GAL4-VP16, to a larger extent than those from the region A and D mutants. We propose that both terminal regions (A and D) play important roles in RNA polymerase assembly, while the central portion (regions B and C) is involved in activated transcription.

journal_name

Curr Genet

journal_title

Current genetics

authors

Mitobe J,Mitsuzawa H,Ishihama A

doi

10.1007/s002940100203

subject

Has Abstract

pub_date

2001-06-01 00:00:00

pages

210-21

issue

4

eissn

0172-8083

issn

1432-0983

journal_volume

39

pub_type

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